BOSTON, CAMBRIDGE, Mass. & CHEVY CHASE, Md.—A team of researchers led by scientists at Massachusetts Eye and Ear, the Broad Institute of MIT and Harvard and the Howard Hughes Medical Institute (HHMI) recently shared their development of a CRISPR/Cas9 genome-editing therapy to prevent hearing loss in a mouse model of human genetic progressive deafness, announcing as well that results of their work had been published in Nature in a paper titled “Treatment of autosomal dominant hearing loss by in vivo delivery of genome editing agents.”

As the researchers noted, hearing loss is the most common form of sensory loss in humans, and nearly half of these cases of deafness have an underlying genetic cause. In their work, they delivered their CRISPR/Cas9 treatment directly to ear hair cells (which are the sound-sensing cells of the inner ear) of mice, successfully preventing hearing loss in those animals.

One of the root causes of genetically based hearing loss is a single-letter mutation in a gene called TMC1 that causes hair cells to produce a toxic protein that accumulates and kills the hair cells over time, particularly during youth—this is a characteristic in both humans and mice.

The purpose of the therapeutic approach was to disrupt a mutation that would otherwise cause the cells to die. The scientists say that their work represents the first time that a genome-editing protein has been ferried directly into the relevant cells to halt progression of genetic hearing loss. Delivering the Cas9 protein itself locally, instead of DNA elements that the cell can use to build Cas9, improved the DNA specificity and potential safety of the treatment, according to the team.

“We set out to develop a genome-editing strategy to try to address this genetic hearing loss by disrupting the underlying genetic variant,” said co-senior author David Liu, the Richard Merkin Professor, director of the Merkin Institute of Transformative Technologies in Healthcare and core institute member at the Broad Institute, as well as a professor of chemistry and chemical biology at Harvard University and an HHMI investigator. “A lot of additional work is needed before this strategy might inform the development of a therapy for humans, but at this stage, we’re delighted and excited that the treatment preserved some hearing in the animal model.”

One of the challenges is that, because the mutated TMC1 gene only differs from its healthy counterpart by a single DNA letter, any Cas9 therapy needs to target the mutated gene with extremely high precision, lest the Cas9 protein cut and disable the functional copy of the gene instead. The team built on prior work published in 2014 to address this issue of editing precision. Typically when CRISPR/Cas9 is used for gene editing, researchers insert the DNA encoding the Cas9 complex into a cell, then allow the cell use its own machinery to produce the gene-editing arsenal.

But the researchers has previously shown that if the Cas9 gene-editing complex itself was delivered directly into a cell, packaged inside an envelope of lipids, the editing was much more precise.

“The strategy we used was particularly efficient in targeting dominant genetic hearing loss,” said co-senior author Zheng-Yi Chen, an associate professor at Massachusetts Eye and Ear. “In humans, dominant hearing loss generally manifests as late-onset and progressive, therefore providing us with a precious time window for intervention. The therapeutic effect through local inner ear delivery also presents a major advantage in reducing potential risks.”

In the untreated mouse model, the animals experience hearing loss by four weeks of age and profound deafness at eight. The treated mice maintained a substantial amount of their hearing compared to the untreated mice. Physiological measurements showed that the hair cells survived at a higher rate in the treated cochlea; genetic sequencing showed that among the edited cells, the mutated copy of Tmc1 had successfully been disrupted 94 percent of the time, and the wild-type allele had only been hit 6 percent of the time. At eight weeks, treated mice also retained their instinctive physical “startle” response to sudden loud sound, while the untreated mice did not respond.

“This is an exciting study that demonstrates the feasibility of a DNA-free, virus-free genome-editing strategy for a type of autosomal dominant hearing loss characterized by progressive hair cell loss,” said Tina Stankovic, an associate professor at Massachusetts Eye and Ear who was not involved with the study. “Augmenting the toolbox to treat genetic deafness is of major significance.”

Getting a bit more technical, as the team’s Nature paper notes in the abstract, “We designed and validated, both in vitro and in primary fibroblasts, genome editing agents that preferentially disrupt the dominant deafness-associated allele in the Tmc1 (transmembrane channel-like gene family 1) Beethoven (Bth) mouse model, even though the mutant Tmc1Bth allele differs from the wild-type allele at only a single base pair. Injection of Cas9–guide RNA-lipid complexes targeting the Tmc1Bth allele into the cochlea of neonatal Tmc1Bth/+ mice substantially reduced progressive hearing loss. We observed higher hair cell survival rates and lower auditory brainstem response thresholds in injected ears than in uninjected ears or ears injected with control complexes that targeted an unrelated gene. Enhanced acoustic startle responses were observed among injected compared to uninjected Tmc1Bth/+ mice. These findings suggest that protein-RNA complex delivery of target gene-disrupting agents in vivo is a potential strategy for the treatment of some types of autosomal-dominant hearing loss.”

Next steps include testing the therapy in larger animal models of progressive hearing loss with a genetic basis. While much work remains before human experiments would move forward, Liu believes that if the treatment pans out, it will work best when administered during childhood, noting: “The conventional thinking in the field is that once you’ve lost your hair cells, it’s difficult to get them back.”